Infection by the Human Immunodeficiency Virus (HIV) causes depletion of CD4 T cells and leads to a state of immuno-suppression. The viral replication requires stable integration of its genome into the human chromosome, yet the natural process of HIV infection leads to the accumulation of a large quantity of unintegrated viral DNA in the body. In spite of clear demonstration of transcriptional activity from non-integrated viral DNA in vitro, little progress has been made in understanding regulation of this early process and its direct role in the HIV life cycle. As such, pathogenic potential of non- integrated viral DNA in HIV infection is not well understood. The long-term objectives of this application are to understand the molecular mechanisms that regulate pre- integration transcription and the contribution of this early process in viral pathogenesis. The specific aims of this proposal are to determine the viral DNA template regulating pre-integration transcription, and the involvement of HIV Tat in the initiation of pre- integration transcription. The experiment methods to achieve these specific aims are to use an HIV integrase negative virus and a novel indicator cell that we have recently constructed to characterize pre-integration transcription. It is expected that data acquired from these studies will make help to gain insight into circumstances where non-integrated viral DNA is capable of producing viral factors, enhancing viral replication and contributing to viral pathogenesis. The proposed research will help to understand potential pathogenic role of non-integrated HIV DNA in disease progression. Additionally, non-integrated viral DNA could serve as clinical markers for disease progression in certain situation. [unreadable] [unreadable] [unreadable]